Systems and Methods for Cultivating, Harvesting and Processing Biomass

ABSTRACT

Combining controlled open-ocean iron enrichment with a system for collecting the ensuing biological growth can lead to a fundamental shift towards using marine biomass feedstock for large-scale global biodiesel production. The literature review reveals that open-ocean enrichment effectively reduces both the atmospheric carbon dioxide partial pressure and ocean acidity. A semi-closed ocean system is provided that allows for the efficient cultivation and harvesting of a high tonnage biomass feedstock generated by iron fertilization. The concept methodically capitalizes on the ocean&#39;s free nutrients, kinetic/potential energy, and expansive surface area to ensure that the mass, energy, and cost balance equations favor our system while taking care to preserve the ocean&#39;s ecosystem. The system is modular, portable, easily scalable system, and minimizes waste. In addition to the above benefits, our concept allows continued adherence to the NEPA and London Protocol by culling the biomass produced by fixing carbon dioxide and limiting iron exposure to the vessel&#39;s interior.

CROSS-REFERENCE TO RELATED APPLICATION

This application is a divisional application of U.S. patent applicationSer. No. 13/375,220, filed Feb. 16, 2012, which is a 371 application ofPCT Application No. PCT/US2010/037037, filed Jun. 2, 2010, which claimsthe benefit of U.S. Provisional Application No. 61/183,447, filed Jun.2, 2009, and U.S. Provisional Application No. 61/263,340, filed Nov. 20,2009, all of which are incorporated herein by reference in theirentirety.

BACKGROUND OF THE INVENTION

Unfortunately, the United States' economy and lifestyle are based onhaving a steady and inexpensive petroleum hydrocarbon supply availablefor import from foreign sources. Consequently, this petroleum supply anddemand dynamic will siphon more and more hard capital and jobs from thedomestic economy as the century progresses due to diminishing crude oilreserves. Earth's crude oil reserves are large but finite in volume.Many scientists predict that crude oil production from proven reserveswill peak within this century. The decline in production will adverselyaffect the United States' oil-based economy and lifestyle. Due toimpending fossil fuel supply issues this century, the U.S. continues toshift significant resources to identify realistic and economicallyviable avenues to produce a domestic renewable hydrocarbon equivalent.

Initially, U.S. scientists focused on using fatty acids from terrestrialplants and animals to make biodiesel fuels. The biodiesel produced is acompatible substitute for fossil fuel diesel but not close to being arobust inexpensive equivalent. The main issues with using terrestrialplant and animal fatty acids are:

-   -   These feedstocks are expensive and account for as much as 80% of        the fuel's cost. According to Anthony Radich and Rudy Pruszko,        the best way to reduce renewable diesel and jet fuel production        cost is by lowering feedstock cost. See, e.g., Pruszko, R        (2007), “Alternative Feedstocks and Biodiesel Production”,        Presented at the Practical Biodiesel Blueprint Conference,        Radich, A. (2004); “Biodiesel Performance, Costs, and Use”        Energy Information Administration, p 4, Website        http://www.eia.doe.gov/oiaf/analysispaper/biodiesel/pdf/biodiesel.pdf,        which is hereby incorporated by reference in its entirety.    -   There are limited quantities of tallow, waste vegetable oil        (WVO), and soybeans.    -   Terrestrial feedstocks cultivation compete with other needs for        land and water resources.    -   Several plant feedstocks are also foods thus increasing the        demand on a limited supply.

In order to address these issues, renewable fuel research has focused onmicroalgae cultivated in freshwater ponds and bioreactors because oftheir high yield per hectare as well as their high lipid and proteincomposition relative to other photoautotrophs. See, e.g., Ingole B S,Parulekar A H, (1995) “Biochemical-Composition Of Antarctic ZooplanktonFrom The Indian-Ocean Sector” Indian Journal Of Marine Sciences 24(2):73-76, which is hereby incorporated by reference in its entirety. Theproblems with using microalgae produced from land based aquacultures,ponds, and bioreactors are: high capital cost, high fresh waterconsumption, generates waste streams, not portable, and not easilyscalable.

Therefore, a need exists for improved systems and methods forcultivating biomass. A further need exists for improved systems andmethods for harvesting and processing biomass.

SUMMARY OF THE INVENTION

The invention provides for terrestrial microalgae aquacultures andbioreactors that can be used to cultivate microalgae in open waterenvironments to generate a steady and inexpensive feedstock. Thesystems, devices, and methods described herein can be used to producerenewable fuels, natural and engineered proteins, and forbioremediation.

The invention provides for improved growth and production of biomass ina vessel situated in open-water environment by retaining iron and ironcompounds within the vessel.

The vessels can include semi-permeable membranes, magnetic fields,buoyancy controlled components, and/or baffles that allow for efficientand cost-effective growth and production of biomass. The vessels can bedesigned to withstand harsh environmental conditions, while allowingexchange of nutrients through the semi-permeable walls of the vessel.

In accordance with an aspect of the invention, a vessel can comprise asemi-permeable exterior wall, wherein the semi-permeable exterior wallselectively retains a microorganism over water-soluble nutrients; and abaffle for directing flow within the vessel in a recirculating patternwhen fluid is passed through the vessel.

In accordance with another aspect of the invention, a vessel maycomprise a buoyant top; a buoyancy-controlled base; and a semi-permeablemesh material connecting the buoyant top to the buoyancy-controlledbase, wherein the semi-permeable mesh material selectively retains amicroorganism over water-soluble nutrients.

A system for producing biomass may be provided in accordance withanother aspect of the invention. The system may include a vessel in anaquatic environment capable of growing a microorganism in an interiorregion enclosed therein, wherein the vessel has a semi-permeablematerial capable of retaining the microorganism in the interior regionwhile allowing fluid from the aquatic environment to flow through. Thesystem may also include a processing platform in fluid communicationwith the interior region of the vessel and configured to harvest themicroorganism from the vessel.

One or more vessels can be secured to existing open-water structures,such as oil-well platforms, barges and boats, and shoreline. A mobile orfixed processing platform can be connected to the vessels to allow forprocessing of the biomass products.

An aspect of the invention provides a method for producing biomass. Themethod for producing biomass can include growing a microorganism in avessel comprising a semi-permeable membrane in an aquatic environment;and retaining iron compounds within the vessel.

Other goals and advantages of the invention will be further appreciatedand understood when considered in conjunction with the followingdescription and accompanying drawings. While the following descriptionmay contain specific details describing particular embodiments of theinvention, this should not be construed as limitations to the scope ofthe invention but rather as an exemplification of preferableembodiments. For each aspect of the invention, many variations arepossible as suggested herein that are known to those of ordinary skillin the art. A variety of changes and modifications can be made withinthe scope of the invention without departing from the spirit thereof.

INCORPORATION BY REFERENCE

All publications and patent applications mentioned in this specificationare herein incorporated by reference to the same extent as if eachindividual publication or patent application was specifically andindividually indicated to be incorporated by reference.

BRIEF DESCRIPTION OF THE DRAWINGS

A better understanding of many of the features and advantages of theinvention will be obtained by reference to the following detaileddescription that sets forth illustrative embodiments, in which many ofthe principles of the invention are utilized, and the accompanyingdrawings of which:

FIG. 1 shows a vessel design that incorporates a magnetic core to retainferromagnetic iron sulfate and a porous material exterior to (a) retainthe oxidized iron, other trace metals, and the induced microalgae and(b) allow hydrophilic micronutrients to enter the vessel.

FIG. 2A shows a vessel with a compression mechanism, which permitsbioreactor volume reduction in an extended state.

FIG. 2B shows the vessel shown in FIG. 2A in a compressed state.

FIG. 3A shows a vessel having a fiberglass composite top and based witha fiberglass mesh material that allows nutrients to be exchanged, whileretaining marine microalgae within.

FIG. 3B shows an exposed electromagnetic solenoid and carbon dioxidebubbling from the fiberglass composite base.

FIG. 3C shows iron powder applied to a solenoid that can provide aninduced magnetic field to orient and retain iron within the vessel.

FIG. 4A shows a top view of a two-liter vessel with a fiberglass fabricexterior and a centrally located electromagnet.

FIG. 4B shows a close-up top view of the two-liter vessel after threedays.

FIG. 5 shows a vessel design incorporating baffles to circulate themicroalgae through the vessel.

FIG. 6 shows an offshore aquaculture implementation to generaterenewable hydrocarbon fuels and proteins.

FIG. 7 shows an implementation of vessels for nitrogen and phosphorousmicronutrient bioremediation along rivers and tributaries prior toreaching coastal estuaries and deltas.

FIG. 8 is a diagram depicting microalgae processing into hydrocarbonfuels and purified proteins.

FIG. 9 shows a vessel base with an electromagnetic solenoid forproducing a magnetic field and fluidic connections for providing astream of carbon dioxide.

FIG. 10A shows a vessel for plankton cultivation having a collapsiblemagnetic coil.

FIG. 10B shows top view of a biomass processing system having aplurality of vessels connected to a processing platform that allows forfarming of open water, e.g., the ocean.

FIG. 11 shows a table with calculations for determining the number ofvessels required to produce 5 million gallons per year at various finalconcentrations of microalgae.

FIG. 12A shows an electromagnetic core design having a singleelectromagnet that can produce a constant and large enoughelectromagnetic field to ensure that trace metals remain in the vesselthat can compress and extend with the vessel.

FIG. 12B shows an electromagnetic core having a first electromagnetpositioned at the perimeter of the vessel and a second electromagnetpositioned at the center of the vessel to promote mixing and/or movementof trace metals between the electromagnets.

FIG. 13 shows equations relating to the scaling of biomass production.

FIG. 14 shows a table showing system scaling based on final vesselplankton concentration and the US daily petroleum distillateconsumption.

DETAILED DESCRIPTION OF THE INVENTION

While preferable embodiments of the invention have been shown anddescribed herein, it will be obvious to those skilled in the art thatsuch embodiments are provided by way of example only. Numerousvariations, changes, and substitutions will now occur to those skilledin the art without departing from the invention. It should be understoodthat various alternatives to the embodiments of the invention describedherein may be employed in practicing the invention.

The invention provides systems and methods for cultivating, harvesting,and processing biomass. Various aspects of the invention describedherein may be applied to any of the particular applications set forthbelow or for any other types of feedstock. The invention may be appliedas a standalone system or method, or as part of a biofuel or biomassproduction system or method. It shall be understood that differentaspects of the invention can be appreciated individually, collectively,or in combination with each other.

The invention provides for various designs, methods, and systems thatallow for the goals set forth herein for the growth of one or moremicrobial organism and the various beneficial effects of the growth,including the production of a renewable fuel, remediation, andproduction of proteins. The invention provides for design features thatwill improve the specific growth rates, product density/specificity, andprevent trace metals from being diluted by ocean or open-water currents.These features and combinations of these features have not beenincorporated in prior ocean enclosure systems. See, e.g., Brockmann U H,Dahl E, Kuiper J, et al. (1983). “The Concept Of Poser (PlanktonObservation With Simultaneous Enclosures In Rosfjorden)” MarineEcology-Progress Series 14 (1): 1-8; S. Takeda, J. N., C. S. Wong, F. A.Whitney, W. K. Johnson, and T. J. Soutar. (1999). “Application ofopen-ocean enclosures to study the control of biological carbon dioxidepump in the subarctic North Pacific Ocean.” Proceedings of the 2ndInternational Symposium Carbon Dioxide in the Oceans: 583-586; and Wang,J. K. (2003). “Conceptual Design of a Microalgae-based recirculatingoyster and shrimp system. Aquacultural Engineering 28: 37-46, which arehereby incorporated by reference in their entirety.

FIG. 1 shows an example of an open-water enclosure in accordance with anembodiment of the invention. A collapsible porous material 100 may beprovided that may allow sea water to pass through but does not allowbiomass to diffuse through. The collapsible porous material may be meshnetting. Open-water enclosures can utilize mesh netting with small poresizes. For example, mesh netting may utilize pore sizes less than 30 μm.Mesh of this size may allow continuous nutrient replenishment throughthe mixing by ocean currents, which captures the microalgae product. Themesh netting can be of any pore size and/or material that allows forselection against loss of trace metals and biomass while allowingtransport of other nutrients. The selection may be charge, size, and/orhydophobicity/hydrophilicity based.

An open water enclosure may include a magnetic core 102. In someembodiments, the magnetic core may be a collapsible magnetic coil.

A large buoyant top 104 to support the structure can be incorporated.The top structure may or may not enclose the vessel. In someembodiments, the top structure may be an air filled hollow cap. In someembodiments, the structure may include a buoyancy controlled bottom 106.Optionally, the vessel may include one or more reinforcement beams 108a, 108 b, 108 c.

Open ocean/water cultivation can allow for following advantages:

-   -   Free nutrients,    -   Free kinetic energy for mixing,    -   Free organism cooling and hydration,    -   The ocean's vast surface area eliminates natural scaling        limitations, and/or    -   Portability and ease of implementation in most ocean        environments.

As a result of these advantages, the products generated, includingrenewable fuels, can be produced for significantly less cost. Ahydrocarbon equivalent can also be produced at a significantly lowercost by various processes, including hydrotreating an oil extract.

Hydrotreating oil extracts allows for the production of a renewablediesel that can:

-   -   Be a biofuel indistinguishable from petroleum diesel,    -   Be a drop-in replacement for or can be blended to any proportion        with traditional diesel,    -   Be used in any proportion in today's infrastructure from        pipelines and storage to gas pumps and automobiles,    -   Be used in any proportion,    -   Has excellent stability and is not oxygenated,    -   Offer superior cold flow properties making it more suitable for        very cold climate conditions,    -   Have higher energy content per volume compared to Biodiesel,        and/or    -   Offer lower fossil energy requirements and reduction of Green        House Gases and NOx emissions.

Iron Fertilization

In some embodiments of the invention, vessels may be used to capture allor substantially all biological growth induced by iron fertilization,which can be accomplished by the exposure of the organism to tracemetals that are confined within the vessel. Metals within the vessel mayinclude reduced iron. In order to prevent the reduced iron fromdiffusing outside the vessel, the vessels may include an electromagneticor permanent magnetic core designed to minimize reduced iron dilution byocean currents or other currents. Other nonmagnetic metals and oxidizediron may be captured by a charged fiberglass material that may allowhydrophilic nutrients through but retain the microalgae and traceelement. The material need not be made of fiberglass and can be formedfrom other materials that provide similar selection characteristics.

Microalgae may require iron to assist in converting carbon dioxide intosugars using light energy from the sun. The oceans iron concentrationsare generally well below the levels required to induce exponentialgrowth in marine algae. As a result, marine microalgae are by and largein a stationary phase until storms or other natural events transportiron from land sources to coastal waters. Trace metals, especially iron,are high beneficial for inducing microalgae growth in oceanenvironments. See, e.g., Martin J H, Coale K H, et al (1994), “TestingThe Iron Hypothesis In Ecosystems Of The Equatorial Pacific-Ocean”Nature 371 (6493): 123-129; Coale, K H, and et al. (1996). “A massivephytoplankton bloom induced by an ecosystem-scale iron fertilizationexperiment in the equatorial Pacific Ocean.” Nature 383: 495-501; Zhou GJ, Bi Y H, Zhao X M, et al. (2009) “Algal Growth Potential And NutrientLimitation In Spring In Three-Gorges Reservoir, China” FreseniusEnvironmental Bulletin 18(9): 1642-1647; Marchetti A, Varela D E, LanceV P, et al. (2010) “Iron and silicic acid effects on phytoplanktonproductivity, diversity, and chemical composition in the centralequatorial Pacific Ocean” Limnology and Oceanography 55(1):11-29; Coale,K H (1991). “Effects of iron, manganese, copper, and zinc enrichments onproductivity and biomass in the subarctic Pacific” Limnology andOceanography 36: 1851-1864; and Coale K H, Johnson K S, Chavez F P, etal. (2004). “Southern Ocean Iron Enrichment Experiment: Carbon Cyclingin High- and Low-Si Waters.” Science 304(5669): 408-414 (herein afterthe “2004 Coale paper”), which are hereby incorporated by reference intheir entirety. Artificially elevating iron concentrations in the openocean is one key to inducing marine microalgae growth as a feedstock forindustrial scale renewable diesel and jet fuel production.

In the 2004 Coale paper, microalgae growth was induced by adding ironsulfate to 0.7 nM in a 225 km² area in the arctic polar front zone ofthe southern seas. The microalgae eventually covered approximate 2400km² after 20 days of growth. Assuming the microalgae had at least adepth of 10 m and a density of at least 1 μg/L, this can yield a wetmicroalgae biomass of approximately 26.5 tons. In some embodiments,algae growth can be tailored toward biological sequestration of carbondioxide. In other embodiments, algae growth can be tailored toward theproduction of a renewable fuel feedstock.

In order to control the cultivation and harvesting of the microalgaeinduced by iron fertilization, several vessels can be utilized with thefollowing design goals:

-   -   To contain microalgae and iron while allowing hydrophilic        nutrients to pass between the vessel and the surrounding        environment.    -   To concentrate microalgae 15× prior to harvesting in order to        reduce the volume of seawater to be processed and discarded and        in turn to reduce the energy requirements to process the        microalgae.

In some embodiments, the goal can be to capture all biological growthinduced by iron fertilization, which can be accomplished by allowing theorganism to be exposed to the trace metals only in the confines of thevessel.

Vessel Design

A semi-closed plankton cultivation and processing system, including avessel, is shown in FIG. 1 and FIG. 10A. The system can include acontrol system (not shown) for analyzing and processing, among otherthings, environmental conditions, nutrient parameters and vessel status.This vessel has several innovations that improve the specific growthrates, product density/specificity, contain (or capture) organisms to begrown, and prevent trace metals from being diluted by ocean currents.

In some embodiments, the vessel may have a substantially cylindricalshape. For example, a cross-sectional area of the vessel may have acircular shape. In other embodiments, the vessel may have any othershape, which may include a prism with a square cross sectional shape,diamond cross-sectional shape, rectangular cross-sectional shape,triangular cross-sectional shape, pentagonal cross-sectional shape,hexagonal cross-sectional shape, octagonal cross-sectional shape, orelliptical cross-sectional shape. In some embodiments, the vesselcross-sectional shape may correspond to a shape of a top apparatusand/or bottom base.

The vessel may include a collapsible porous material 100 that may allowsea or ocean water to pass through without allowing biomass to diffusethrough. In some embodiments, the collapsible porous material may be amesh netting. The collapsible porous material may be formed from afabric or textile, such as a cotton canvas material. In someembodiments, the porous material may be formed of a synthetic,non-synthetic, or blended fiber. The porous material may be formed ofone or more materials. The porous material may be a filtration membrane.The porous material may be flexible enough to allow stirring by oceancurrents. In some embodiments, the porous material may be porous enoughto allow sea/ocean/surrounding water to diffuse through. The materialmay also be dense enough to contain product within the vessel, by makingits diffusion timescale sufficiently large. Various embodiments of theporous material are discussed further below.

The collapsible porous material may surround a magnetic core 102. Insome embodiments, the magnetic core may be one or more collapsiblemagnetic coil. The magnetic core may include any magnetic material.Preferably, the magnetic material may have a shape or configuration thatmay enable it to collapse in a vertical direction. For example, it mayinclude a coil, telescoping features, sliding features, foldingfeatures, accordion-type features, small loose components, or otherconfigurations that enable collapsing. Various embodiments of themagnetic core are discussed further below.

The collapsible porous material may be provided between a top apparatus104 and a bottom base 106 of the vessel. One or more reinforcing beams108 a, 108 b, 108 c may be provided between the top apparatus and thebottom base.

The exterior vessel design incorporates innovative ideas to reduceenergy consumption and operating cost requirements. One feature is abuoyant top apparatus to support the structure. In some embodiments, thetop apparatus may be filed with air. In other embodiments, the top maybe filled with another gas or material that may have less density thanwater. In some embodiments, the top may be hollow or may include poresthat may trap air or other gases. The top apparatus may be formed of anymaterial or have any configuration that may allow the top apparatus tofloat on the ocean's surface.

The concept significantly improves the functionality of the vessel byadding a buoyancy controlled base as show in FIG. 1 and FIG. 10A. FIG.2A shows that when the base 200 is filled with water, it expands thecultivation volume/depth of the vessel. In some embodiments, the vesselmay be expanded so that the base reaches of a depth of 30 m. In otherembodiments, when the vessel is expanded, the base may be at any depth,which may include but is not limited to about, up to about, or greaterthan about 100 m, 70 m, 50 m, 40 m, 35 m, 30 m, 25 m, 20 m, 15 m, 10 m,7 m, or 5 m.

Once the biomass has reached a predetermined density, air is pumped intothe base 200 causing the cultivation volume/depth to decrease as thebase rises. The base may be brought closer to a top 202 of the vessel.As the vessel approaches the compressed state in FIG. 2B, the productconcentration increases by as much as 15×. In some embodiments, theconcentration can increase about, up to about, or greater than about 5,10, 15, 20, 25, 30×. Concentration can be achieved by flow or efflux ofwater, such as seawater, through a semi-permeable meshing 204. Thiscritical processing step significantly reduces the volume of seawaterand in turn reduces the energy requirements for the pumps and dewateringcentrifuges during biomass processing.

Semi-Permeable Meshing

A variety of materials may be chosen for the flexible porous materialthat allows seawater to pass freely but impedes plankton (or otherbiomass or fuel) diffusion out of the vessel. In some embodiments, thevessel includes a porous material that may be constructed of Nitexnetting with a pore size less than 20 μm, a fiberglass upper portion,and a lower buoyancy control base. The netting can be protected fromrips by something similar to a high density cotton canvas exterior.Polyamine and polyethylene netting material with pores less than 20 μmcan be used. Other materials can also be used. One, two, three, four, ormore layers of flexible porous material may be provided. In someembodiments, the layers may be formed of the same material, while inother embodiments, the layers may be formed of different materials. Insome instances, the layers may have the same pore size, while in otherinstances, the layers may have different pore sizes.

Functionally, the mesh may allow for transport of water and other smallmolecules through the mesh and retention of organisms to be grown, suchas those described herein. The mesh may be a semi-permeable exteriorwall that selectively retains the organisms over water-solublenutrients. For example, the wall can retain more organisms thanwater-soluble nutrients. For example, the wall can retain 2×, 3×, 5×,10×, 15×, 20×, 30×, 40×, 50×, 70×, 100×, 200×, 500×, 1000× or moreorganisms than water-soluble nutrients by mass, concentration or volume.The prototype can be tested in a 3 million gallon closed waterenvironment at Moss Landing Commercial Park (MLCP) for construction andmaterial quality. A plankton concentration time course can be conductedby inoculating the vessel interior with plankton so that a initialvessel concentration of 1 g/L is achieved and measure for Chlorophyll a(Chl a) inside and outside the vessel.

Not only for efficiency concerns, but also for environmental concerns,the semi-permeable material can be selected to minimize the transport ofbiological mass to the surrounding environment. The correct combinationof materials and construction can be achieved when the baseline Chl alevels in the seawater are maintained outside the vessel whilemaintaining close to the original 1 g/L plankton concentration insidethe vessel.

Magnetic Core Design

Trace metals, especially iron, are the key to inducing plankton growthin ocean environments. Since the goal is to capture all biologicalgrowth induced by iron fertilization, it may be desirable for organismexposure to the trace metals to occur only in the confines of thevessel. In order to prevent the trace metals from diffusing outside thevessel, an electromagnetic core designed to minimize trace metaldilution by ocean currents is provided. Para- and ferromagneticparticles may be retained within the vessel.

Examples of potential electromagnetic cores are depicted FIG. 12A andFIG. 12B. The one electromagnetic field in FIG. 12A is constant andlarge enough to insure that the trace metals, especially iron, remain inthe vessel. A one-coil design may include a magnetic coil that maycompress and extend with a vessel. It may be possible to tailor tracemetal concentrations to the prevailing organism growth conditions byvarying the electromagnetic field intensity as a form of controlledrelease. An alternative is the design in FIG. 12B that incorporates anelectromagnetic field at the perimeter and in the center of the vessel.A two-coil design may be provided, which may compress and extend withthe vessel. Alternating the electromagnetic field between the twoelectromagnets promotes trace metal mixing within the vessel whilepreventing them from being diluted.

One or more electromagnetic coils may be in electrical communicationwith a power source. The power source may be part of the vessel or maybe external to the vessel.

As previously described, a magnetic core may include a magnet orelectromagnet that may be compressible or collapsible. The magnetic corecan preferably be compressible or collapsible in a vertical direction.

The magnetic core may be enclosed by the vessel. For example, themagnetic core may be enclosed by a semi-permeable or porous material.The magnetic core may be entirely closed by the semi-permeable or porousmaterial. In some embodiments, the magnetic core may be at leastpartially enclosed by the semi-permeable or porous material.

Exemplary Vessel Designs

One embodiment of a vessel is shown in FIG. 1 and FIGS. 2A and 2B. Avessel can have one or more of the following design features:

-   -   Variable and/or collapsible electromagnetic core to retain the        reduced iron sulfate as shown in FIG. 1.    -   Constant replenishing of hydrophilic nutrients.    -   Constant dilution of waste organic acids excreted by the        microalgae.    -   Culture never dehydrates or over heats.    -   Concentrates organism by reducing vessel volume as shown in FIG.        2.

The vessel, as shown in FIG. 1, may have a maximal volume of 9.4 millionliters, a diameter of 20 meters, and a height that can be adjusted from2 meters to 30 meters. The vessel may contain a compressible orcollapsible magnetic or electromagnetic core 102 that can facilitateretention of trace metals, e.g., magnetic trace metals, or iron. Thevessel may be cylindrical in shape or any other suitable shape. Anydiscussion herein of a cylindrical shape may apply to any other shapeand vice versa.

The circumferential walls 100 of the vessel may be made of a meshmaterial that allows selective transport of nutrients, e.g., nitrogen,phosphorous, water, waste products, trace metals, and biomass. The meshmaterial may be made of fiberglass with a particular pore size. In someembodiments, the porous material may be made of a plastic, a metal, aglass, an organic material, or any combination thereof that has thedesired selectivity. In some embodiments of the meshes or nettings, orporous materials described herein can be fabricated using fiberglassfabric, carbon fiber fabric, polyethylene, polyvinylacetate, orhydrophilic polymer. These materials can be woven. In some embodimentsof the invention, these materials can be charged, e.g., fiberglassfabric. Some other examples of materials include, e.g., a durable andinexpensive high density cotton canvas material, a filtration membrane,a metallic sheet, any woven hydrophilic fabric or polymer (e.g., wovenpolypropylene, woven polyethylene), or membrane with a selected poresize. Selected pore sizes include any pore size of about or up to about1, 5, 10, 15, 20, 25, 30, 40, 50, 75, 100, 150, 200, 250, 300, 500,1000, 2000, 3000, or 5000 microns. In some embodiments, the pore sizemay be variable or uniform across the surface of the mesh material orinto the mesh material. Alternatively, pore size may vary within themesh material. The pores may be evenly distributed over the meshmaterial, may be grouped into clusters, or may have any otherdistribution over the mesh material.

As stated previously, the selectivity can be achieved by pore size,charge, hydrophilicity/hydrophobicity, magnetism, or any combinationthereof. In some embodiments, the selectivity is obtained by providingfor a material that has a relatively greater diffusion time for thebiomass and other materials to be retained as compared to materials thatare to be exchanged with the surrounding environment.

The top 104 of the vessel and the base 106 of the vessel can beconnected in a variety of manners. In some embodiments, the top and baseare connected only by the cylindrical walls 100 of the vessel. Aninterior region of the vessel may be enclosed by the cylindrical wallsand/or the top and base. The cylindrical walls may or may not beflexible and/or collapsible. In some embodiments, the cylindrical wallsare rigid. The top and base of the vessel can be movable with respect toone another. They may be movable in a vertical direction. They may ormay not be movable in a horizontal direction. In one example, whenorganisms are contained within the vessel, the top and base of thevessel may be brought into closer proximity with one another. The topand base may be brought into closer proximity while the walls arecollapsed. The walls may optionally be collapsed without expandinghorizontally. In some embodiments, the walls may be collapsed whilemaintaining a substantially same cross-sectional area enclosed therein.This may reduce the volume within the vessel. Reducing the volume of thevessel may result in increasing concentration of the organism therein.

In other embodiments, the top and base are connected by the cylindricalwalls and one or more reinforcing beams 108 a, 108 b, 108 c. In someembodiments, one, two, three, four, five, six, seven, eight, or morereinforcing beams may be provided. The reinforcing beams can provide fora rigid connection between top and base, while the cylindrical walls canremain flexible. The one or more reinforcing beams can limit themmobility of the base with respect to the top. For example, thereinforcing beam can limit the mobility of the base with respect to thetop in a vertical direction. The base may move along the rigidreinforcing beams (as shown in FIG. 2) and/or the rigid cylindricalwalls such that the base moves vertically and is prevented fromtranslating horizontally away from the top of the vessel. In someembodiments, the reinforcing beams may remain fixed with respect to thetop of the vessel, and the base may slide with respect to thereinforcing beams. Alternatively, the reinforcing beams may be fixedwith respect to the base of vessel, and the reinforcing beams may slidewith respect to the top of the vessel. In other embodiments thereinforcing beams may be movable with respect to both the top and bottomof the vessel. In some embodiments, the top and/or bottom may includeone or more vertical channel or passage that may be capable of slidingwith respect to the reinforcing beam. In other embodiments, the topand/or bottom of the vessel may be provided between the reinforcingbeams. The top and/or bottom of the vessel may be configured to remainat a substantially fixed horizontal/lateral position with respect to thereinforcing beams.

The position of the vessel base relative to the top of the vessel can becontrolled in a variety of manners, e.g., buoyancy and/or mechanicallycontrolled. In some embodiments, the base and the top of the vessel maybe buoyancy controlled. The top of the vessel may be an air filledhollow cap and the base of the vessel may be negatively buoyant. Thebase may also have one or more compartments that allow for buoyancycontrol. In some embodiments, the compartments may be or may containbladders that can hold air or another gas. Once the bladders are filledwith gas, the base may become positively buoyant and cause the base torise toward the top of the vessel. The bladders can be filled in amanner such that the rate of vessel volume reduction or expansion iscontrolled. This may be important to increase the selectivity of porouscylindrical walls, e.g., to selectively allow transport of water andother undesired materials across the porous walls and retain iron andbiomass. The bladders may be in communication with one or more gassource. For example, one or more hose or channel may connect thebladders to a gas source. In some embodiments, a pump, or positivepressure source may be provided to force gas into the bladders to raisethe base. The bladders may also be in communication with one or more gasvent, when it is desirable to lower the base. The bladders may ventdirectly to the surrounding water, or may be connected to a remoteventing location via a hose or channel. In some embodiments, the remoteventing location may be the same as, or different from, the gas source.

In some embodiments, a vessel base may be raised and/or lowered via oneor more mechanical actuator. For example, the base and/or top may moverelative to a reinforcing beam via an actuator. Examples of actuatorsmay include but are not limited to, motors, solenoids, linear actuators,pneumatic actuators, hydraulic actuators, electric actuators,piezoelectric actuators, or magnets. Actuators may cause the base and/orreinforcing beam to move based on a signal received from a controlsystem. The actuators may or may not be connected to a power source.

FIGS. 3A, 3B, 3C, 4A, and 4B show an example of a vessel describedherein. The electromagnetic vessel can be referred to as the IronFertilization Vessel (IFV). A 1^(st) generation 2 L prototype IFV (FIG.3A) has been built and has successfully been shown to confine growth toIVF's interior (FIG. 4A and 4B), to limit iron exposure to the IVF'sinterior by inducing an electromagnetic field (FIG. 3C), to concentratethe culture 3× prior to harvesting by compressing IFV's volume, and toblow carbon dioxide gas from the base (FIG. 3B). FIG. 4A shows the 2 Lvessel prior to incubation in water (t=0). FIG. 4B shows the 2 L vesselafter three days of incubation in water. The grey matter is the reducediron powder attached to the magnet. Red matter on the walls of thefiberglass fabric is mostly oxidized iron retained by the fiberglassfabric. The oily substance present throughout the interior of the vesselshown in FIG. 4B may indicate the presence and/or growth of microalgae.

The 1^(st) generation IFV prototype is equipped with a two ply of an 8.5oz 2×2 twill weave fiberglass mesh, a fiberglass composite top, andbuoyancy controlled base, as shown in FIG. 3A. A fiberglass fabric maybe selected as the mesh material because it is a strong, flexible, andhydrophilic material that allows nutrients to be replenished andsimultaneously contain the microalgae inside the vessel. The fiberglasscomposite buoyancy control base can have two internal compartments. Theupper compartment can be used to blow carbon dioxide from the base. Thelower compartment can be filled with seawater during the growth phase tomaximize surface area in which nutrients can diffuse across. The lowercompartment can be filled with air during the harvesting phase toconcentrate the algae by decreasing the IFV's volume prior to processingand/or harvesting the marine algae.

The vessel can include an electromagnetic core designed to retain theiron additions to the IFV, as shown in FIG. 3C. The electromagnetic coilin the 1^(st) generation 2 L prototype consisted of two wires wrappedaround an iron core with opposite polarities. A timing circuit can allowthe current to alternate between the two wires, thus changing thedirection of the magnetic field. The wave function in the magnetic fieldcauses the iron to oscillate between the two ends of the solenoidallowing it to mix with the aqueous media and algae inside the vessel.

The electromagnetic field strength is approximated by Ampere's Law asB=μ_(o)ηI, where μ₀ is the permeability of the core, η is the number ofturns per unit length, and I is the current. The B field strength isdirectly proportional to the current. The current can be regulated bychanging the resistance in the circuit. Once the magnetic field has beeninduced, its magnitude is determined by a calibrated Hall Effect sensor.The magnetic field exerting at least 6.4 mT can retain reduced ironinside the vessel 6.4 mT, as shown in FIG. 3C.

The invention also provides for designs that can be utilized tostreamline the vessel's functionality and operation by increasingcirculation and/or mixing within a vessel, as shown in FIG. 5. Forexample, the vessel can be designed to mimic fresh water raceway pondfunctionality but in a vertical fashion. Increased circulation can allowfor better mixing of nutrients and allow for desired or selectedexposure of the microalgae to sunlight. The mixing and flow dynamics canbe controlled by the design of the vessel dimensions and shape. Baffleswithin the vessel can be used to control fluid velocity and volumetricflow rates for liquid flowing in a recirculating pattern within thevessel and for liquids flowing through the vessel. The current in thesurrounding environment can also be utilized to prevent fouling of themesh materials by directed current flow across the mesh material or bypowering a mechanical cleaning device. In some embodiments, the currentcan be used to generate power, which can be utilized by the vesselitself in any form, or by the processing platform.

To afford protection from the elements, the vessel, including thesemi-permeable walls, can be rigid. In some embodiments of theinvention, one wall can be rigid and the other can be flexible. Thevessel can be designed to be resistant to damage by weather, current, orany large objects in the surrounding environment. The vessel can bedesigned to be rigid and protective, while not substantially restrictingflow into and out of the vessel from the surrounding environment. Insome embodiments, the vessel can be buoyancy controlled to allow thevessel to be submersed during inclement weather. Buoyancy control can beachieved by the top portion, the base, or any combination thereof.

Loss of microorganism and other nutrients through the upstream orcurrent-facing side of the vessel is less of a concern than loss throughthe down-stream facing portion. In some embodiments, the up stream orcurrent-facing side of the vessel can have a first pore size and thedown-stream facing side portion can have pores of a second size that aresmaller than the first size.

As shown in FIG. 5, a permanent magnet that spans the width of thevessel may be incorporated to increase the retentions time that thereduced iron sulfate (C) remains in the upper region of the vessel. Themagnet can have a minimum field strength of 6.4 mT. In addition to acomponent for retaining iron, the vessel may also include components forretaining other nutrients. For example, the vessel may includemechanisms to concentrate nitrates and/or phosphates. The mechanism mayinclude chromatography components, ion-exchange based materials, e.g.,ion-exchange columns, and/or affinity based materials, e.g., affinitycolumns Any of the vessels described herein may have components forconcentration and/or retention of one or more nutrients, e.g., iron,nitrate, and/or phosphate compounds.

Stainless steel sieved gate (shown as dashed lines between C and A inFIG. 5) with a pore size less than the organism. In some embodiments,the gate is enclosed by the vessel. For example, the gate may beentirely enclosed by the vessel, or at least partially enclosed by thevessel. The gate may be movable between a blocking position and an openposition. The gate may be lifted or moved out of a blocking position toan open position to allow the organism to circulate. The gate may belowered or put in a blocking position to concentrate and harvest theorganism (A). FIG. 5 shows the gate (dashed line) in a blockingposition.

FIG. 5 indicates a deficient nutrient feed point (B). Feeds that are lowin concentration in the surrounding environment can be added at point B.The deficient nutrients that can be fed to the vessel include anynutrient discussed herein. In some embodiments, the nutrients includeiron, phosphate, and/or nitrate compounds. The iron can be fed as aniron compound, such as iron sulfate, or iron can be fed to the vessel aspart of a biodegradable polymer or material that releases iron overtime, as discussed herein. The biodegradable polymer or material canalso include other nutrients, such as nitrate compounds and/or phosphatecompounds. Nitrates can also be fed in the form of ammonium, ammoniumferrous(II) sulfate (magnetic), or ammonium bicarbonate. Nitrates andother nutrients can also be sourced from waste water, secondary wastewater, run off, chicken feed, agricultural waste, or any low-costnutrient source and then fed to the vessel. The nutrient feed can becontrolled automatically or manually. The nutrient feeding may becontrolled based on the concentration of the nutrient in the vessel, thegrowth rate and/or the concentration of the organism. A nutrient feedingcomponent for feeding one or more nutrients can be included in any ofthe vessels described herein.

Uses the current's kinetic energy to thoroughly mix the micronutrientsand the microalgae. The mixing of nutrients and algae can be achieved bybaffles within the vessel that direct the fluid in a recirculatingpattern. The vessel may be positioned within a flowing current. In FIG.5, current flows into the vessel at the right-hand side (D right) andexits the vessel at the left-hand side (D left). The movement from rightto left forces circulation within the vessel in the direction indicatedby the arrows, which forms a recirculating pattern. The circulation canbe created by a Venturi effect caused by the flux of fluid through thereactor from the upstream portion of the vessel to the downstreamportion of the vessel. The amount of current flow used for circulationcan be selected in a variety of manners, e.g., by altering the exposedsurface area on the right hand side of the vessel and/or the surfacearea on the left-hand side D. In this configuration, the vessel has anupstream, or current-facing side and a downstream or a side that is notfacing the current. If the current of the surrounding environment isfixed, the vessel may be fixed in a proper orientation. If the currentis not fixed, then the directionality of the vessel may be controlledbased on the current's direction. The control of the vessel'sorientation can be automatic or manual.

As described above, orientation of the vessel relative to the current inthe surrounding environment can plan an important factor in determiningthe circulation rate within the vessel. To account for this, the vesselcan be designed such that the orientation of the vessel with respect tothe direction of current flow can be controlled. A self-orientingmechanism capable of orienting the direction of the vessel can beprovided. Mechanical features, such as vane-like features, can be usedto self-correct or self-orient the direction of the vessel such that adesired flow of water through the vessel is achieved. For example, oneor more fin, protrusion, channel, flap, or shaped feature can beprovided for the vessel. A self-orienting mechanism can be provided in astationary position relative to the vessel, or can be movable relativeto the vessel.

In some embodiments, the vessel orientation with respect to the currentis such that maximal flow through the vessel is achieved. In otherembodiments, the vessel orientation can be such that flow through thevessel is lower than the maximal flow through the vessel. For example,if maximal flow is achieved by placing the incoming mesh side the vesselperpendicular to the flow, a lesser amount of flow can be achieved byplacing the incoming mesh side at an orientation that is notperpendicular to current flow in the surrounding environment.

Uses the current's kinetic energy to concentrate the microalgae. Oncethe sieve gate shown in FIG. 5 is placed in a blocking position, thecirculation, as described above, can be utilized to concentrate themicroalgae against the sieve gate.

All the microalgae spend the same cumulative time in the sun exposurezone (between A and B in FIG. 5). The amount of time spent exposed tothe sun can be controlled based on the circulation rate through thevessel and the cross-sectional area of the channels that allow exposureto the sun relative to the cross-sectional area of the other channels inthe vessel.

The recirculation caused by the flux of water through the vesselmaintains a constant microalgae density throughout thecirculating/recirculating portion of the vessel.

The vessel shown in FIG. 5 can be designed for high Reynolds and Pécletnumber to insure it is in the convection regime for consistent nutrientand organism density.

The pivot point (G) shown in FIG. 5 can control the incoming watervelocity. As described above, circulation may be controlled by a varietyof manners. Here, an incoming water gate can control or restrict therate of water entering the vessel.

If necessary, the vessel percolates or sparges carbon dioxide from thebase (F) shown in FIG. 5 in an effort to achieve higher microalgaedensities.

A hydrophilic, charged, porous material (D) shown in FIG. 5 can allowenvironmental micronutrients and waste organic acids to cross freely butcontain the microalgae. This can be achieved by selecting an appropriatepore size, e.g., less than about 5, 10, 15, 20, 30, 50, 100, or 150 μmpore size (or any other pore size described herein).

Organisms and Metabolic Engineering

While microalgae and plankton have been referred to as organisms to begrown within the vessels, a variety of organisms can be grown in thevessels described herein. These organisms can include plankton, diatoms,algae, phytoplankton, and zoo plankton. The organisms to be grown can beselected based on geographic considerations. The organism can be anyautotrophic or photoautotrophic organism. In some embodiments, theorganisms grown within the vessels are more than one type of organism.For example, symbiotic organisms can be grown in conjunction with eachother, or one organism may be grown during a first phase and a secondorganism may be grown during a second phase. In some embodiments, theorganisms grown in the vessel can include an organism that performsnitrogen fixation. Nitrogen-fixing organisms can be grown with algae orany other organism in a symbiotic relationship. Examples of organismsthat perform nitrogen fixation include Richella intracellularis,nitrogen-fixing blue-green algae, nitrogen-fixing cyanobacteria, andTrichodesmium.

The organisms, e.g., microalgae, utilized herein can be metabolicallyengineered for the efficient conversion of the nutrients to furtherincrease the microalgae growth rate, improve product yields, decreasevessel requirements, and maximize the overall system productivity. Themicroalgae can be engineered to: increase fatty acid content andrenewable fuels, e.g., biodiesel, productivity, manufacture industrialenzymes, synthesize personal care/medicinal proteins, and/or manufacturespecialized fuels, e.g., jet fuel.

Isolate Local Wild Type Strains and Seed Culture Growth

In some embodiments, a large microalgae seed culture can be inoculatedat the time the iron is added to the vessel to accelerate microalgaegrowth relative to diatoms. Microalgae can be the dominant species grownand harvested due to its higher initial concentration. Local wild typestrains, which may be preferred, can be used such that new species arenot introduced into the local environment. Local microalgae and diatomstrains can be characterized for specific growth and nutrientrequirements. As well, environmental parameters can be determined suchthat microalgae or diatom growth can be selectively induced.

Processing Platform

The vessel products (e.g., biomass produced by growth of or productionby organisms grown within the vessels) can be harvested using aprocessing platform 1010 (see FIG. 10B). The processing platform can bea mobile unit that can be connected to the vessels 1012 describedherein. The processing platform may be in fluid communication with aninterior region of one or more vessel. In some embodiments, theprocessing platform can be connected to the vessels via a connector 104.In some embodiments, a connector may be a hose, pipe, or channel. Insome embodiments, the processing platform and/or vessels can include apump, a positive pressure source, or a negative pressure source, totransfer organisms within the vessel to and/or from the processingplatform. The processing platform can optionally connect to each vesseldirectly. Alternatively, the processing platform can connect to one ormore hubs, which can connect to one or more vessel. A processingplatform can be connected to any number of vessels, including but notlimited to one or more, two or more, three or more, four or more, fiveor more, six or more, seven or more, eight or more, nine or more, ten ormore, twelve or more, fifteen or more, twenty or more, thirty or more,forty or more, or fifty or more vessels.

The processing platform can be located on a rig or an oil tanker. Theprocessing platform can be located on a buoyant or floating support. Theprocessing platform can also be located on land, which may be in closeproximity to a body of water. For example, the processing platform canbe located on a shoreline.

The processing platform can process organisms in real-time, after acertain period of time, or periodically. Lipids contained withinorganisms can be recovered and processed into biodiesel. Carbohydratesand proteins contained within the organisms can be collected and used inproducts for human or animal consumption. Seawater can be rejected andreturned to the ocean. The processing platform can produce substantiallyno toxic or hazardous waste streams.

Methods

Microalgae Physiology:

The target culture density can be controlled. In some embodiments, thetarget control density is in the range of about 1 μg/L to 100 g/L, 100μg/L to 50 g/L, or 1 to 10 g/L.

Nutrient uptake rates can be measured and accounted for to sustainselected growth rates or biomass production rates.

The waste organic acid accumulation rate can be measured and accountedfor. The rate of waste organic acids transport out of the vessel throughthe permeable mesh membrane may also be controlled by adjusting thecirculation rate, or by selecting preferred mesh materials.

The iron concentration can be measured and controlled. Ironconcentration may be a limiting factor for growth or may provide a meansto control growth rate.

The concentration of silica, nitrogen, phosphorous, and carbon nutrientscan also be measured and controlled. In some embodiments, thesenutrients can pass through the membrane at a higher rate than iron andbiomass. The concentration of silica, nitrogen, phosphorous, and carbonnutrients can affect growth and productivity of the microalgae and alower non-inhibitory concentration for these compounds can be determined

The iron feed profile can be selected for a number of parameters. Theiron feed profile can be adjusted based on the growth of the organismwithin the vessel, or the iron feed profile can be adjusted to maintainconstant or varying iron concentration.

In some embodiments, various strains may have different specificity fordifferent iron compounds. The iron compound used can be selected forefficient growth of the organism of choice of vice-versa, the organismcan be selected for efficient growth using a predetermined ironcompound.

The production of renewable fuel and protein yield per gram iron can bemeasured and optimized based on other parameters described herein.

Various environmental parameters that naturally select for and maximizespecific growth rate can be monitored and accounted for.

Iron Fertilization Vessel

Iron containment and the concentration of iron within the vessel may bemeasured using optical density or chelating/pH measurements on samplestaken inside and outside the vessel.

Microalgae containment may be measured using optical densitymeasurements from samples taken inside and outside the vessel duringgrowth experiments.

Membrane material investigation can be selected such that it is durableand hydrophilic.

Vessel Automation, Sampling and Processing

In some embodiments, LCMS can be used to measure water samples upstreamand downstream of the vessels. Other parameters that can be measuredinclude optical density (to determine the concentration of the culture),iron concentration in the culture, and culture pH. These procedures,including sampling and processing of the biomass using the methodsdescribed herein, can be automated.

Organism Selection and Preparation

The following provides one example of organism selection andpreparation. First, local sea water can be retrieved, which may includeseawater from the Sea Cliff Beach Pier in the Monterey Bay, to be usedas a first source of seawater. Low silicate sea water can be used as asecond source of seawater. Iron may be added to each sea water type tobring its iron concentration to 5 nM. 1 mM sodium bicarbonate can beadded to each sea water type as a substitute for CO₂ gas. Both sea waterstocks can be filtered with a 0.2 μm hydrophilic membrane. 15 g agar canbe added to 500 mL of each sea water stock. The agar may be dissolved bymicrowaving until everything it is melted. The sea water stocks can beremoved from the microwave and shaken. The petri dishes can be filled toabout half full. The petri dishes can be let to solidify and storedupside down in a cold room.

Next, unfiltered local marine water can be streaked on both plate types.They may be incubated on the bench in the presence of a constant fullspectrum light source. From the filtered local marine water plates, anisolated diatom colony may be picked and streaked on new local marinewater plates. From the low silicate plates, an isolated microalgaecolony may be picked and streaked new low silicate plates. These platesmay be incubated until single colonies grow. A single diatom ormicroalgae colony can be picked from a plate and a sterile test tubefilled with 5 ml of filtered local marine water stock and with 5 nM Feand 1 mM NaHCO₃ (Solution C) can be inoculated. This may be incubated ina shaker with a full spectrum light source present until the culturereach 1 OD at 600 nm. Next, a sterile 500 mL flask filled with 50 mL ofSolution C and all 5 mL of the first generation seed culture can beinoculated. This can be incubated in a shaker in the presence of a fullspectrum light source present until it reaches 1 OD at 600 nm. Thesecondary seed culture should either be (i) used to inoculate a closedwater time-course experiment and/or (ii) split into a master stock foreach specimen.

Electromagnetic Intensity

The minimum electromagnetic field required to bind the bulk of the tracemetals can be determined In a clear fresh water tank with constantagitation, the electromagnetic potential can be maximized to bind allthe trace metals to the sealed coil. Starting from the maximumpotential, the electromagnetic potential may be incrementally decreaseduntil trace metals start to release from the coil allows for thedetermination of the minimum electromagnetic field required to bind thetrace metals. After each field adjustment, a spectrophotometermeasurement at 595 nm can be taken. This can continue until thespectrophotometer readings increase 10% over or under the baseline.

Iron Compound Selection

Introducing iron into an ocean environment typically induces the largestbiological response of all the trace metals. Iron sulfate has been thesubstrate used in most open water fertilization experiments. However, itmay not be the best choice because it induces the growth of bothplankton and diatoms. Based on the multiple composition profiles in theliterature, plankton is preferred over diatoms because it has asignificantly larger lipid and protein composition. The diatomcomposition is skewed towards ash/fertilizer products. Therefore, aniron compound that is preferentially selected by plankton may bepreferred.

A selected iron compound can be chosen by performing laboratory growthtime course experiments using different iron compounds and 0.2 μmfiltered seawater. The idea is to maximize plankton specific growthwhile minimizing the diatoms' at the same time. Also fed-batchintroduction of the iron compound may promote plankton growth overdiatoms.

Iron compounds can be selected for specificity for diatoms andmicroalgae. As well, the amount of iron can be selected such that aminimal amount of iron is used to maintain a desired growth rate. Ironconcentrations can be about or up to about, or at least about 5.00,2.50, 1.25, 0.63, 0.31, 0.16, 0.08, 0.04, 0.02, 0.01, or 0.005 nM iron.

In some embodiments of the invention, the iron compound is formed withina biodegradable material. The biodegradable material can be any suitablepolymer that allows for desired release of iron compound at the properconcentration. The biodegradable material can allow for time-release ofthe iron compound and can also allow for the iron compound to beencapsulated within a large particle that will not pass through thesemi-permeable walls of the vessel. Furthermore, encapsulation of theiron compound can reduce or prevent oxidation of the iron compound.Oxidation of the iron compound can change the magnetic properties of theiron.

The following is an exemplary procedure to test for desired ironconcentration:

First, make a stock local sea water solution with 1 mM sodiumbicarbonate (Solution A), a carbon source substitute for the CO₂ gas.Add iron sulfate to 250 mL of the Solution A to achieve a concentrationof 5 nM (Solution B). Achieve the above growth media iron concentrationsby serially diluting Solution A with Solution B. Add iron containsolutions to test tubes and inoculate with the appropriate strains.Incubate the test tubes in the presence of a continuous full spectrumlight source. From the above experiment, the highest iron concentrationlevel that will induce microalgae and impede diatom exponential growth(lowest concentration level diatoms) can be determined

Maximize Microalgae/Plankton Specific Growth Rate

Iron is one of the first limiting compounds in ocean environments (See,e.g., Scharek R, Van Leeuwe M A, De Baar H J W (1997). “Responses ofSouthern Ocean Phytoplankton To The Addition Of Trace Metals” Deep-SeaResearch II Vol. 44 (1-2): 209-227). Once iron is supplemented, the nextlimiting trace metal or nutrient can be identified. Utilizing microbialphysiology, plankton specific growth rate can be maximized bysupplementing quickly depleted nutrients and/or trace metals to thevessel. In essence, the vessel can be used as an open-water fermenter toachieve high growth rates and plankton density. After maximizing growthby regulating extracellular factors, the plankton can be metabolicallyengineered for the efficient conversion of the nutrients to furtherincrease the plankton growth rate, improve product yields, decreasevessel requirements, and maximize the overall system productivity.

Time-courses can be conducted to track the depletion rates of importantnutrients and trace metals by microalgae, diatoms and other organisms.Depleted nutrients can be fed at a rate that sufficiently alleviates theobserved rate-limiting factor that occurs during the exponential growthphase. At that point, a new rate limiting compound can be identified andfed accordingly. This process of identifying and feeding deficientnutrient can repeat itself until the predetermined growth rate isachieved or all the environmental parameter modification options havebeen exhausted.

In a 20-55 L closed water environment, add filter sterilized local seawater with 1 mM sodium bicarbonate or blow carbon dioxide from the baseof the 2 L prototype as shown in FIG. 9. At time zero, inoculate thevessel interior with the 50 mL seed culture and start feeding iron andother depleted nutrients. The feed rate of the iron and depletednutrients will be ramped up according to their utilization profile fromprevious experiments and the current instantaneous growth rate. Takesamples at 2 hr intervals until the culture reaches the stationaryphase. At each sampling point measure the pH, the optical density at 600nm, and the mass density inside and outside the vessel. Analyze thesamples for micronutrient concentrations and their depletion rates,especially phosphates, nitrates and bicarbonates. Analyzing for nitratescan give some insight in to whether this concept can effectively be usedto remove excess nitrates that have a hand in inducing Harmful AlgaeBlooms (HABs). Analyzing Bicarbonate concentrations may give someinsight as to how large a role this concept may play in recyclinggreenhouse gases and global warming. With total microalgae amounts, howmuch carbon dioxide and nitrates are being removed from the environmentand converted to microalgae that will be processed into renewable fuels,protein extract, and glycerol can be determined

Additional Technologies

In conjunction with the systems, methods, and devices described herein,techniques and scientific knowledge from the following areas can also beharnessed:

-   -   1. Fatty acid extraction and transesterification    -   2. Glycerol purification and recovery to sell to pharmaceutical        and personal care product manufacturers.    -   3. Bulk protein, amino acids, and carbohydrate recovery and        packaging.

Microalgae Bioprocessing

One example of a conversion process from microalgae growth to therenewable fuel and protein products, described herein, is shown in FIG.8. The biomass grown in the vessels described herein are initiallyscreen filtered prior to dewatering with a screw press. The microalgaeoils are dried prior to being fed to the delayed coking unit and thenhydrotreated. During the early implementation of this process, theextracted microalgae oil can be sold to fossil fuel refiners to becoprocessed with their incoming crude oil.

During an implementation, the biomass from the vessels may be screenfiltered and then dewatered with a screw press. The dried biomass mayundergo protein separation. Then solar drying may occur to for purifiedproteins.

Delayed Coking

Delayed coking is a thermal process which has two majorreactions—thermal cracking and polymerization. Thermal cracking is themechanism through which molecules of high molecular weight in the feedstock are decomposed into smaller, lighter molecules that arefractionated into their end products. Polymerization is a reactionthrough which many small hydrocarbon molecules are combined to form asingle large “coke” molecule of high molecular weight. A typical cokehas 100 to 200 carbon molecules.

The main objective of the delayed coking unit is to convert microalgaeoil to lighter products of higher value and to produce a coke product.In some embodiments, fresh microalgae feed is preheated through a heatexchange system prior to entering the bottom of the coker fractionatingtower. The fresh microalgae feed is mixed with recycle from the unitbefore being pumped through two fired heaters. The effluent from theheaters then enters the bottom of the coking drums where the gaseousproducts pass out the top and the liquid soaks in the drum until itcracks into lighter products that will exit the top of the drum or formscoke. The liquid and gaseous products resulting from the thermalcracking are separated into the desired products by fractionation in adistillation tower before being deoxygenated via hydrotreating. The cokeproduct in the coke drum is removed batchwise from the drums aftercooling.

Hydrotreating

Conventional hydrotreating technology of microalgae oil extract producesa high quality product that is compatible with existing fuelinfrastructure. Hydrotreating deoxygenates microalgae feedstock byadding hydrogen to produce a highly-stable renewable diesel fuel with ahigher cetane value, lower cloud point and lower emissions thanbiodiesel and traditional petrodiesel.

Facilities/Location

The vessels can be implemented at sites that are near seawater, rivers,estuaries, oceans, lakes, or any body of water. In some embodiments, thesite is near rail service to facilitate transportation of goods.

The growth parameters can be optimized and the microalgae's metabolicpathways can be engineered to achieve a significantly higher specificgrowth rate. Increases in the microalgae's specific growth rate, theinitial culture density (P_(i)) and final culture density (P_(f)) canincrease the overall system productivity and lower the total vesselrequirement.

As shown in FIG. 11, a final microalgae concentration can be at leastabout, up to about, or about 25 g/L, thus requiring only 16 vessels toproduce 5M gallons of fuel per year. Sixteen vessels can utilize orrequire only 1.25 acres of ocean surface area.

Advantages

The invention provides for a systems, devices, and methods that allowfor the growth, concentration, and harvesting of mass marine biomassinduced by iron fertilization for commercial biofuel, e.g., biodiesel,production. The benefits of ocean and open-water farming are numerousand far outweigh any benefits of any land based algae efforts.

The devices and methods describe herein have comparatively low feedstockproduction costs. The reduced production and distribution costs arelargely due to the following advantages: (i) very little overhead isrequired to run and maintain farms in the open ocean, (ii) productionvessels can be quickly fabricated and deployed off any coast, close toend users particularly in major urban areas and near transit arterieswith very little initial capital, (iii) the ocean has a free andabundant supply of all the required nutrients to produce renewablefuels, except the very inexpensive iron compounds, (iv) feedstockproduction is highly scalable, and (v) virtually no waste streams aregenerated, which would require additional disposal costs and fees.

Capital and Operational Costs

The systems described herein have a number of advantages. The vesselconstruction is extremely low compared to bioreactors. The vessels aremodular and portable. The vessels do not require highly skilled labor tooperate and optimize it like bioreactors. As a result, they can beestablished in most coastal regions of the world; moreover, they do notrequire laborers with specialized skills.

The vessels do not utilize limited resources like land and fresh water.It reduces rents/mortgages requirements due to the minimum land use. Thecultivation, harvesting, and microalgae fuel processing stages can bedone at sea thus eliminating the cost of leasing land. Most landrequirements can be for storage and management office space. Theinvention does not compete with other developed land uses such asfarming for food or residential/commercial developments. In fact, theonly resource that is required is the widely abundant iron. Thisminimizes or reduces chemical purchasing requirements to iron anddownstream processing chemicals such as ethanol/methanol and basecatalyst. The majority of the cultivation nutrients are provided andcontinuously replenished by the ocean for free, thus lowering the rawmaterial expenses.

The magnetic iron retention and/or mixing mechanism increase the overallyield and productivity of the system thus minimizing the vesselrequirements and capital investment. The ocean provides the kineticenergy required to keep the system well mixed for free thus furtherreducing energy costs and requirements.

Additionally, no waste streams are produced. There are no waste streamsbecause (1) the supernatant is returned to the ocean (during growth andduring dewatering/harvesting), (2) the biomass lipids are converted tobiodiesel, (3) and the carbohydrate/protein extract are packaged forhuman/animal consumption.

The use of buoyancy as the concentrating mechanism to reduce vesselvolume prior to processing in order to minimize the volume that needs tobe pumped and centrifuged provides a significant advantage. Usingbuoyancy is a less energy intensive method other mechanical methods.

In comparison to open-ocean iron fertilization where biomass grown isnot harvested, the invention provides for product revenue streams thatare generated from (1) converting the microalgae lipids to renewablefuels, and (2) extracting the algal carbohydrate/protein forhuman/animal consumption.

Also, tax credits from both carbon sequestration and biodiesel/renewablefuel production can be obtained.

Manufacturing Benefit

In addition to the cost benefits, there are several manufacturingbenefits to take into account. The system that we described above isconstructed of modular components that can be easily fabricated out ofcomposite materials, e.g., fiberglass or carbon fiber compositematerials, once the molds for the parts have been made. Because of themodular composite components, these vessels can easily be repaired,replaced, and relocated. When the vessel is displaced by a storm, theycan easily be located if the system components are both filled with airmaking them buoyant and a locating beacon is installed. Finally, thesystem is scalable to sizes that are impractical on land due to avariety of reasons, including volume and surface area limitations.

Economic

The invention provides for system, devices, and methods that can createa domestic and worldwide renewable hydrocarbon fuel source, createworldwide distribution, bioprocessing, manufacturing, and engineeringjobs, and can potentially replace the need for all imported diesel fuelproducts. Also, it ensures the US maintains a technological advantage indeveloping and deploying energy technologies.

Engineering

The invention provides for a platform technology that can be used forgenerating medicinal, industrial, and nutritional proteins worldwide.

Military

The invention allows for local fuel production and depots around theworld. This can be useful to military installations that are dispersedthroughout the world, but have access to aqueous environments.

Environmental and Safety Issues

The environmental benefits are straight forward but just as important asthe other benefits. The biomass produced fixes dissolved CO₂, causingsurrounding atmospheric CO₂ pressure to decrease. The ocean waters arealso simultaneously deacidified. The oceans can be deacidified byfixation of dissolved carbonate and bicarbonate

The invention can be used to restore coastal ecosystems throughbioremediation. The invention can be used to remediate coastal estuariesand river deltas by reducing nitrogen and phosphorous concentrations andeliminating unwanted hazardous algae blooms.

Another benefit worth discussing is the ramification and cleanup of anaccidental spill of biodiesel or unprocessed marine biomass. Since thebiomass/biodiesel is biodegradable, it can be either consumed ordegraded by other organisms in an ocean environment over the course ofno more than 17 days.

In some instances, the organisms to be grown can be contained within thevessels. This can reduce the environmental impact of organisms on thesurrounding environment. For example, genetically modified organisms canbe substantially contained within the vessel.

The devices and methods herein can be designed such that compliance withEPA, NEPA, and London Protocol regulations is met. In the open ocean,time courses experiments can be conducted to determine if there is anyperturbation to the surrounding marine ecology and how fast thesurrounding marine environment recovers after cultivating and harvestinglarge quantities of plankton. The invention provides for systems,devices, and methods that allow for farming of the oceans withoutdisrupting ecological systems and continue to comply with NEPA and theLondon Protocol.

Another potential environmental and safety concern is what effect doescreating an artificial magnetic field has on the surrounding ecologicalsystem. Electromagnetic field intensity can be controlled such that theocean habitat's health and the safety of the employees is protected.

Because the plants can be located throughout the world, this canminimize the fuel consumed while distributing fuels to end users.

With all these benefits, the cost and energy balance equations are verymuch in favor of our system.

Quantitative Impact: Scaling Calculation

The following calculations were made to determine the number of vesselsand ocean surface area required to replace the US daily consumption ofpetroleum distillate. First, the time needed for a culture to start atan initial concentration, P_(i), and grow to a final extended vesselconcentration, P_(f), needs to be calculated from Equation 1, shown inFIG. 13. Once the time is calculated the biomass and lipid productionrates per vessel can be determined from equations 2 and 3 shown in FIG.13, respectively.

Since glycerol is replaced by methanol or ethanol during thetriglyceride transesterification, we assumed that the lipid contentweight is approximately equal to the biodiesel product weight. As aresult, the biodiesel production rate per vessel is approximated byequation 4 shown in FIG. 13.

Now that the biodiesel production rate per vessel has been approximated,we can determine the ocean farm size required to displace the petroleumdistillates used by the United States each day by equations 5 and 6shown in FIG. 13.

The results of the above calculations based on literature values for thegrowth rate are listed in the table shown in FIG. 14. In order togreatly reduce the vessel requirement numbers in FIG. 14, we canengineer the vessel and the stains to achieve a significantly higherspecific growth rate, initial culture density (P_(i)) and final culturedensity (P_(f)). Improving the above three factors can increase theoverall system productivity and lower the total vessel requirement. Atthe very least, we believe that this system can replace 25% of thepetroleum fuel consumption in the United States.

Application: Renewable Fuel and Protein Synthesis

In some embodiments of the invention, the vessels are located around oildrilling platforms off the US Pacific and Gulf of Mexico coastlines.Offshore drilling platforms have been in the service of the oil industryfor decades. Once the oil reserves are depleted, these platforms arecapped and left to decay into man-made reefs. These inactive offshoreplatforms dot the Gulf of Mexico coastline without a useful purposeuntil now. Offshore platforms are ideal for mooring aquacultures. FIG. 6displays a possible layout around an out of service drilling platform600. The platform offers a place to harvest and process the microalgaeon site into renewable hydrocarbon fuels, protein supplements, andglycerol which is preferred because it is more efficient to transportliquid fuels. As shown in FIG. 6, a tanker or river barge retrofittedwith the tools necessary for doing the microalgae to renewable fuelsconversion at sea is also a viable option.

In some embodiments, one or more vessel 602 may be provided upstream ofthe platform 600.

Application: Bioremediation of Coastal Estuaries and River Deltas

The devices and methods described herein can be used for remediatingcoastal estuaries and river delta regions, as shown in FIG. 7. Theright-hand side of FIG. 7 shows two potential implementation points ontributaries that release into the Chesapeake Bay estuary. As an example,the vessels 700 can be implemented for nitrogen and phosphorousmicronutrient bioremediation along rivers and tributaries prior toreaching coastal estuaries and river deltas. Microalgae grown in vesselsdescribed herein can be processed into purified proteins and hydrocarbonfuels that may be packaged and sold in the local markets. There may befewer policy hurdles to cross to clean up coastal dead zones and anopportunity to build a positive reputation with the United Statesgovernment and the American people.

Along the coast and tributaries, iron is usually not the limitingnutrient. Once the deficient nutrients for microalgae growth areidentified, they can be fed into the vessel in quantities that correlateto the biomass density. The rate at which nutrients are fed into thevessel may be proportional to the uptake rate determined in a laboratoryclosed water time course experiment. Controlling the harvest rate maydirectly affect the microalgae concentration and indirectly affect thenitrogen and phosphorous uptake rate. As described before, the harvestedmicroalgae can be processed on site into renewable hydrocarbon fuels,concentrated proteins, and glycerol to be sold on the local markets.These alternative profit streams ensure that this remediation endeavoris economically viable. As shown in FIG. 7, the processing plant 702 canbe located on land rather than on water because land is nearby.

Any components, configurations, characteristics, features, or steps asknown in the art may be used in any of the embodiments discussed herein.See, e.g., Kalra, A. and W. S. LLP. (2006). “BiodieselTax Credits”;Walford, L. A. (1958). Living Resources of the Sea: Opportunities forResearch and Expansion. New York, Ronald Press; and Löscher, B. M.(1999). “Relationships among Ni, Cu, Zn, and major nutrients in theSouthern Ocean.” Marine Chemistry 67: 67-102, which are herebyincorporated by reference in their entirety.

It should be understood from the foregoing that, while particularimplementations have been illustrated and described, variousmodifications can be made thereto and are contemplated herein. It isalso not intended that the invention be limited by the specific examplesprovided within the specification. While the invention has beendescribed with reference to the aforementioned specification, thedescriptions and illustrations of the preferable embodiments herein arenot meant to be construed in a limiting sense. Furthermore, it shall beunderstood that all aspects of the invention are not limited to thespecific depictions, configurations or relative proportions set forthherein which depend upon a variety of conditions and variables. Variousmodifications in form and detail of the embodiments of the inventionwill be apparent to a person skilled in the art. It is thereforecontemplated that the invention shall also cover any such modifications,variations and equivalents.

What is claimed is:
 1. A method for producing biomass comprising:growing a microorganism in a vessel comprising a semi-permeable membranein an aquatic environment; and retaining iron compounds within thevessel.
 2. The method of claim 1, wherein the microorganism is at leastone of the following: microalgae, plankton, diatoms, algae,phytoplankton, or zooplankton.
 3. The method of claim 1, wherein thesemi-permeable membrane allows fluid from the aquatic environment topass freely while impeding diffusion of the microorganism out of thevessel.
 4. The method of claim 1, further comprising reducing the volumeof the vessel within the semi-permeable membrane, thereby increasingconcentration of the microorganism therein.
 5. The method of claim 4,wherein the vessel comprises a top and a base, and wherein said reducingthe volume occurs when the top and base are brought into closerproximity to one another.
 6. The method of claim 1, wherein a magnet orelectromagnet is used to retain the iron compounds within the vessel. 7.A system for producing biomass comprising: a vessel in an aquaticenvironment capable of growing a microorganism in an interior regionenclosed therein, wherein the vessel has a semi-permeable materialcapable of retaining the microorganism in the interior region whileallowing fluid from the aquatic environment to flow through; aprocessing platform in fluid communication with the interior region ofthe vessel and configured to harvest the microorganism from the vessel.8. The system of claim 7 wherein the processing platform recovers lipidscontained within the harvested microorganism and processes the lipidsinto biodiesel.
 9. The system of claim 7 wherein the processing platformcollects carbohydrates and proteins contained within the harvestedmicroorganism and processes them into products for human or animalconsumption.